The CRISPR/Cas genome editing system has revolutionized almost every aspect of the life science industry. Until recently, the most used formats for this technology have been plasmids, mRNA, or lentivirus. Each reagent has been successful in its own right, however, each approach has limitations. SygRNATM, the two-part synthetic crRNA and tracrRNA, increases the pace of research, decreases costs, and can be used with Cas9 protein, Cas9 mRNA and Cas9 expressing cells/models.

This webinar discusses the development of the SygRNATM system, protocol optimization, and proposes workflows that enable scientists to quickly incorporate CRISPR technologies into their research.

H&E is the most frequently used stain in histology and is the basis for diagnostics and further selected methods. Thus it is important to have brilliant staining. We will show which minor factors can give a negative effect or spoil the result completely. Tips will be given on improving the sensitivity of the stain. For PAS, we will give advice on how to avoid common errors and always get colourful results.
We will discuss:
•Troubleshooting
•Technical tricks
•How to prevent "critical situations"

In research areas such as academic, biofuel, food and pharmaceutical industries the determination of algal viability, lipid content, and cell concentration is important in the selection, monitoring, and maintenance of algal cultures. Flow cytometry has been shown to be an ideal method to assess health and lipid content of cultures but has been challenging to adopt due to high complexity and cost of traditional technologies. In this webinar, we present novel simplified methods for algal characterization using microcapillary cytometry on either a simple touch screen based cytometer, the Muse® Cell Analyzer, or a higher throughput cytometric platform, the compact Guava® easyCyte platforms. The MilliporeSigma algae kits utilize simple mix and read protocols, dedicated software modules, and provide quick results for the of count and viability measurements or relative lipid content on algae strains. The optimized and dedicated algae kits allow for high precision and comparable results to predicate methods. Data from applications to multiple common algal strains such as Chlorella vulgaris and Chlamydomonas reinhardti under different culture conditions will be presented. Availability of dedicated kits for algae research on simple and easy to use cytometric platforms will empower and enable algae researchers to rapidly select optimal culture conditions and strains for downstream experiments.

Interaction between antigen-specific T cells and antigen presenting cells (APC) cognate ligand involve reorganization of the cytoskeleton and recruitment of adhesive and signaling molecules to the site of intercellular contact. Sustained adhesion of T cells to APCs and formation of the immunological synapse after T cell receptor stimulation are required for the antigen-specific response. One way to measure an immunological synapse is by fluorescently labeling the molecules that have been recruited to the synapse and imaging by fluorescence microscopy. However, immunological synapses are rare and therefore difficult to analyze objectively and statistically by traditional microscopy methods. To overcome these problems, we employed the Amnis brand imaging flow cytometers to objectively collect imagery of large numbers of cells. We report the percentage of T cells involved in an organized immunological synapse, the recruitment of adhesion molecule LFA-1 and signaling molecule Lck to the synaptic complex and subsequent translocation of NFkB from the cytoplasm to the nucleus in the T cell. In this study, Raji B cells loaded with Staphylococcal enterotoxin B (SEB) were incubated with human T cells to create T cell-APC conjugates. Cells were stained in various combinations for CD3, CD19, Actin, LFA-1, Lck and NFkB. Results from the FlowSight and the ImageStream imaging flow cytometers are compared. Using the FlowSight imaging flow cytometer we demonstrate image-based parameters that were used to assess the frequency of conjugates with an organized immunological synapse in an objective and statistically significant manner. Employing the ImageStream imaging flow cytometer we further evaluate the specific location of the adhesion and signaling molecules LFA-1 and Lck within the immunological synapse complex in T cells and measure the nuclear localization of NFkB in the T cell.

Childhood cancer and pediatric cancer are general terms used to describe a wide range of neoplasms found in children and teenagers. Occurring in approximately 1 in 300 people under the age of 20, compared to 1 in 6 adults, pediatric cancers are more rare than adult cancers. Because less is known about pediatric cancers, diagnosis can be quite challenging for pathologists.

This presentation covers the basic science, as well as facts and statistics about pediatric cancer. We will discuss how the utility of immunohistochemical testing along with the application of novel antibodies can contribute to accurate diagnosis and survival rates of pediatric cancer patients.

The ImageStream® and FlowSight® are multispectral imaging flow cytometers that generates high resolution images of cells at a rate of 1000’s of cells per second. This allows for the rapid acquisition of tens of thousands of images per sample. Using the IDEAS® image analysis software, the system calculates features based not only on fluorescence intensity but the morphology of that fluorescence as well. This novel approach is able to seamlessly combine the quantitative power of flow cytometry with the high content information associated with microscopy. The system can collect data on a wide range of applications including nuclear localization during a signal transduction cascade, measuring colocalization of two probes, or quantify features on the phagocytosed particles in macrophages.

Skin cancer is by far the most prevalent cancer. Each year, approximately 3.4 million people in the US alone are diagnosed with some form of skin cancer. Skin cancer can be highly treatable if it is detected and classified early, and this detection and classification is often aided by immunohistochemistry. This presentation covers many of the basic science, facts, and statistics of skin cancer, as well as the utility of immunohistochemical testing with markers such as S-100, SOX-10, Ber-Ep4, and HHV-8 in the accurate diagnosis and survival rates of skin cancer. Continuing education credits for attending this webinar will be offered through the National Society of Histotechnology

A scientific overview of the portfolio of cell lines University of Cambridge has deposited at ECACC with a focus on the KARPAS 299 and KARPAS 422 cell lines; the HeLa Mitotrap cell lines; and CHO cell lines. We will also provide an overview of the process of partnering with ECACC and Sigma-Aldrich for the storage and distribution of cell lines for research purposes. Topics include: The scientific applications of the cell lines; The types of companies and institutions we license to; Advantages of partnering with culture experts and specialised distributors; Our experience of working with ECACC and Sigma-Aldrich.

A scientific overview of the portfolio of cell lines University of Cambridge has deposited at ECACC with a focus on the KARPAS 299 and KARPAS 422 cell lines; the HeLa Mitotrap cell lines; and CHO cell lines. We will also provide an overview of the process of partnering with ECACC and Sigma-Aldrich for the storage and distribution of cell lines for research purposes. Topics include: The scientific applications of the cell lines; The types of companies and institutions we license to; Advantages of partnering with culture experts and specialised distributors; Our experience of working with ECACC and Sigma-Aldrich.

Colorectal cancer is the third most commonly diagnosed cancer in men and women, and is the second leading over all cause of cancer deaths in the United States. Each year, the CDC reports that approximately 135,000 people are diagnosed with colorectal cancer, and over 50,000 succumb each year to the illness. The National Cancer Institute cites a decline in the mortality rate of colorectal cancer due to advanced screening and early diagnosis. Immunohistochemistry has served as the catalyst for these advancements in colorectal cancer diagnosis. This presentation covers many of the basic science, facts, and statistics of colorectal cancer, as well as the utility of immunohistochemical testing with markers such as cadherin¬17, CDX¬2, SATB2, and the mismatch repair proteins in the accurate diagnosis and survival rates of colorectal cancer.

CRISPR Cas9 nucleases have revolutionized the field of genome editing enabling unprecedented efficiency of gene targeting in a vast array of cell types and organisms. Even with such powerful technology at hand, researchers who are new to the field may find genome modification to be challenging and time-consuming. As CRISPR becomes a focus of the molecular biology research community, MilliporeSigma seeks to share the best approaches learned and methods applied in our years of genome editing experience. Today’s presentation will focus on practical applications of CRISPR for pristine genome editing to achieve knockout as well as specific sequence changes to include donor-mediated snps, reporter-tags and conditional knockouts. Special attention will be paid to design considerations for the donor constructs necessary to achieve specific sequence changes. Finally, the frontiers of CRISPR technology, including synthetic crRNA to fast-track genome editing experiments, whole genome screening and targeted gene activation will be explored.

The new EPA METHOD 325 for sampling Volatile Organic Compounds from Fugitive and Area Sources was promulgated in September of 2015. The petroleum refineries have 2 years to comply with the new ruling. Benzene emissions are the focus of the new ruling, but other VOC’s can be monitored using the same sampling tube. The air samples are collected using thermal desorption tubes that are deployed along fenceline of the property and remain deployed for 14 days. The samples are collected passively, without the using of an air sampling pump. After sampling the diffusive endcap is replaced with a metal storage cap and sent to a laboratory where they’re analyzed by thermal desorption gas chromatography. This is the first of a two series presentation that focuses on the analysis side of the workflow. It provides users the steps required to comply with the new EPA fence line regulations and what is needed to optimize the method for high sample throughput. Refineries and testing laboratories will benefit by gaining valuable information on the optimum techniques to perform this method and what it takes to comply. An introduction to the theory and operation of thermal desorption will also be included.

Gene synthesis has become a quick and cost-effective way to build your DNA construct for a variety of applications - from protein engineering to bio-based circuit and pathway design. This webinar, presented by Sigma-Aldrich and GENEWIZ will explain the process of gene synthesis, how it compares to traditional cloning techniques, and review some key applications researchers are addressing with synthetic DNA.

Gene synthesis has become a quick and cost-effective way to build your DNA construct for a variety of applications - from protein engineering to bio-based circuit and pathway design. This webinar, presented by Sigma-Aldrich and GENEWIZ will explain the process of gene synthesis, how it compares to traditional cloning techniques, and review some key applications researchers are addressing with synthetic DNA.

EnPresso, a proprietary growth system from BioSilta Ltd, applies slow enzymatic glucose release to control the growth and protein synthesis rate in bacteria and yeasts. This high-cell-density growth system can be used in any shaken cultivation device at various scales. In this webinar, Dr. Kaisa Ukkonen and Dr. Antti Vasala will review the basic requirements for successful high-cell-density cultivations and discuss common reasons for failures in E. coli-based protein production, the focus being on the cultivation conditions. In particular, prerequisites for successful scale-up will be presented. The webinar also features examples on the use of EnPresso demonstrating the time and cost savings enabled by this powerful growth system, and presents an interesting alternative protein expression protocol: IPTG-based autoinduction.

Sulfonyl fluorides are privileged protein-reactive functional groups due to their context-specific reactivity with a broad range of amino acid residues (tyrosine, serine, lysine, threonine, cysteine and histidine). As a result, they are ideally suited to a myriad of applications in chemical biology and drug discovery. To highlight the growing utility of sulfonyl fluoride chemical probes, examples will be provided from the literature and work in which Dr. Jones has been involved.

Using 2-dimensional nanosheets in a 3-dimensional world:
At the beginning of its exciting life, graphene was mostly a game for physicists. Then, chemists have learned how to “play” with this unique material by enhancing its processability and versatility. Today it is possible, using covalent or supramolecular chemistry, to tailor graphene into a wide variety of forms ranging from simple, soluble sheets to hierarchical architectures where 2-dimensional (2D) graphene sheets are assembled into three-dimensional (3D) composite materials or foams for applications in electronics, mechanics, energy storage or catalysis.
After graphene, the exfoliation of a wide range of layered materials (BN, MoS2, MoSe2, WS2 etc.) has been demonstrated, featuring complementary conductive, semiconductive and insulating properties, opening the way to produce new composite, 2D meta-materials.
More recently, polymerization of small molecules through directional 2D crystallization or cross-linking of self-assembled monolayers was used to obtain self-standing, robust nanosheets featuring a huge variety of chemical structures.
Using these different approaches there is practically no limit to the number of possible 2D structures inspired to graphene that shall be created.

The Sigma-Aldrich Scientific Seminar channel features scientific presentations from key specialists in analytical chemistry, biology, chemistry and life sciences on the practical and technical aspects of new developments and innovations, to help advance your research.